LOCUS       sequence-344927-        6698 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Akt-STOPS targeted to the nucleus via histone H2A..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6698)
  AUTHORS   Zhou X, Zhong Y, Molinar-Inglis O, Kunkel MT, Chen M, Sun T, Zhang 
            J, Shyy JY, Trejo J, Newton AC, Zhang J
  TITLE     Location-specific inhibition of Akt reveals regulation of mTORC1 
            activity in the nucleus.
  JOURNAL   Nat Commun. 2020 Nov 30;11(1):6088. doi: 10.1038/s41467-020-19937-w.
  PUBMED    33257668
REFERENCE   2  (bases 1 to 6698)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun."; date: "2020-11-30"; volume: "11(1):6088. doi"; pages: " 
            10.1038/s41467-020-19937-w"
COMMENT     Sequence Label: pcDNA3-H2A-Akt-STOPS
FEATURES             Location/Qualifiers
     source          1..6698
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(61..80)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        252..631
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        632..835
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     786..806
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     880..899
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        880..898
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1320..2027
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(1465..1483)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1733..1752
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     promoter        complement(2268..2286)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2269..2286)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2306..2323)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2312..2536
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2582..3010
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2669..2688)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2879..2900
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3019..3039)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3024..3353
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3204..3339
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3266..3285
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3420..4214
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3474..3493)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4084..4103
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4388..4509
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4425..4444)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4479..4498
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4558..4574)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4558..4574)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4571..4593)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4582..4598
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(4606..4612,4613..4630,4631..4636))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4651..4672
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4789..4806)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4960..5548)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5040..5059)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5719..6510,6511..6579))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6342..6361
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6580..6684)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 aagtgccacc tgacgtcgac ggatcgggag atctcccgat cccctatggt gcactctcag
       61 tacaatctgc tctgatgccg catagttaag ccagtatctg ctccctgctt gtgtgttgga
      121 ggtcgctgag tagtgcgcga gcaaaattta agctacaaca aggcaaggct tgaccgacaa
      181 ttgcatgaag aatctgctta gggttaggcg ttttgcgctg cttcgcgatg tacgggccag
      241 atatacgcgt tgacattgat tattgactag ttattaatag taatcaatta cggggtcatt
      301 agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg
      361 ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac
      421 gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt
      481 ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa
      541 atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta
      601 catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acatcaatgg
      661 gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg
      721 gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca actccgcccc
      781 attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca gagctctctg
      841 gctaactaga gaacccactg cttactggct tatcgaaatt aatacgactc actataggga
      901 gacccaagct tgcggccgcc accatgtctg gtcgtggcaa acaaggaggc aaggcccgcg
      961 ccaaggccaa gtcgcgctcg tcccgcgctg gcctccagtt cccggtaggg cgagtgcacc
     1021 gcttgctgcg caaaggcaac tacgcggagc gggtgggggc cggcgcgccc gtctacatgg
     1081 cggcggtcct cgagtacctg accgccgaga tcctggagct ggcgggcaac gcggctcggg
     1141 acaacaagaa gacgcgcatc atccctcgtc acctccagct ggccatccgc aacgacgagg
     1201 aactgaacaa gctgctgggc aaagtcacca tcgcccaggg cggcgttttg cctaacatcc
     1261 aggccgttct gttaccaaag aaaaccgaaa gccacaaagc caaaagcaaa gcggatccca
     1321 tggtgagcaa gggcgaggag gataacatgg ccatcatcaa ggagttcatg cgcttcaagg
     1381 tgcacatgga gggctccgtg aacggccacg agttcgagat cgagggcgag ggcgagggcc
     1441 gcccctacga gggcacccag accgccaagc tgaaggtgac caagggtggc cccctgccct
     1501 tcgcctggga catcctgtcc cctcagttca tgtacggctc caaggcctac gtgaagcacc
     1561 ccgccgacat ccccgactac ttgaagctgt ccttccccga gggcttcaag tgggagcgcg
     1621 tgatgaactt cgaggacggc ggcgtggtga ccgtgaccca ggactcctcc ctgcaggacg
     1681 gcgagttcat ctacaaggtg aagctgcgcg gcaccaactt cccctccgac ggccccgtaa
     1741 tgcagaagaa gaccatgggc tgggaggcct cctccgagcg gatgtacccc gaggacggcg
     1801 ccctgaaggg cgagatcaag cagaggctga agctgaagga cggcggccac tacgacgctg
     1861 aggtcaagac cacctacaag gccaagaagc ccgtgcagct gcccggcgcc tacaacgtca
     1921 acatcaagtt ggacatcacc tcccacaacg aggactacac catcgtggaa cagtacgaac
     1981 gcgccgaggg ccgccactcc accggcggca tggacgagct gtacaagcgc atgcatcctc
     2041 gtccgcgctc gtgcacctgg ccggacccca ggccggagtt tggagtcgac cctcgtccgc
     2101 gctcgtgcac ctggccggac cccaggccgg agtttggagg taccggcggc agcgagctcc
     2161 ctcgtccgcg ctcgtgcacc tggccggacc ccaggccgga gttttaagaa ttctgcagat
     2221 atccatcaca ctggcggccg ctcgagcatg catctagagg gccctattct atagtgtcac
     2281 ctaaatgcta gagctcgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt
     2341 gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc
     2401 taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt
     2461 ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca tgctggggat
     2521 gcggtgggct ctatggcttc tgaggcggaa agaaccagct ggggctctag ggggtatccc
     2581 cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
     2641 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
     2701 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
     2761 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg
     2821 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt
     2881 ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta
     2941 taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt
     3001 aacgcgaatt aattctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc
     3061 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt
     3121 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     3181 tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc
     3241 cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc tctgcctctg
     3301 agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctcc
     3361 cgggagcttg tatatccatt ttcggatctg atcaagagac aggatgagga tcgtttcgca
     3421 tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag aggctattcg
     3481 gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc cggctgtcag
     3541 cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc
     3601 aggacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc
     3661 tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg ccggggcagg
     3721 atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct gatgcaatgc
     3781 ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg aaacatcgca
     3841 tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat ctggacgaag
     3901 agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgcgc atgcccgacg
     3961 gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg gtggaaaatg
     4021 gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca
     4081 tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct gaccgcttcc
     4141 tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat cgccttcttg
     4201 acgagttctt ctgagcggga ctctggggtt cgaaatgacc gaccaagcga cgcccaacct
     4261 gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt
     4321 tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc
     4381 ccaccccaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa
     4441 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa
     4501 tgtatcttat catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc
     4561 atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg
     4621 aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt
     4681 gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg
     4741 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga
     4801 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat
     4861 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
     4921 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     4981 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     5041 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     5101 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     5161 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     5221 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     5281 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     5341 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     5401 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     5461 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     5521 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     5581 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat
     5641 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga
     5701 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg
     5761 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga
     5821 gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc
     5881 agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac
     5941 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc
     6001 agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc
     6061 gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc
     6121 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt
     6181 ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc
     6241 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg
     6301 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag
     6361 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat
     6421 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc
     6481 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa
     6541 aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta
     6601 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa
     6661 aaataaacaa ataggggttc cgcgcacatt tccccgaa
//
